Condensation products of glyoxylic acid esters with aminothiols

ABSTRACT

THE INVENTION RELATES TO A THERAPEUTICAL COMPOSITION USEFUL IN PARTICULAR FOR THE PREVENTION AND TREATMENT OF INJURIES DUE TO IONIZING RADIATIONS. SAID COMPOSITION COMPRISES, AS ACTIVE INGREDIENT, A COMPOUND RESPONDING TO EITHER ONE OF THE FOLLOWING FORMULAE:   R&#39;&#39;-CH(-NH2)-CH-S-CH(-COO-R)-OH OR   2-(R-OOC-),4-R&#39;&#39;-THIAZOLIDINE   IN WHICH R IS A STRAIGHT- OR BRANCHED-CHAIN ALKYL GROUP HAVING 1-18 CARBON ATOMS AND R&#39;&#39; IS HYDROGEN, -COOH OR -COOR&#34; IN WHICH R&#34; IS A STRAIGHT- OR BRANCED-CHAIN ALKYL GROUP HAVING 1-18 CARBON ATOMS. THE INVENTION RELATES ALSO TO A PROCESS FOR THE PREPARATION OF THE ACTIVE INGREDIENT OF SAID COMPOSITION AND TO THE NEW COMPOUNDS OF FORMULA A OR B.

United States Patent 3,790,623 CONDENSATION PRODUCTS OF GLYOXYLIC ACID ESTERS WITH AMINOTHIOLS Jean-Pierre Fourneau, Paris, France, assignor to Laboratoires Houde, Paris, France No Drawing. Filed Dec. 16, 1970, Ser. No. 98,921 Claims priority, applicgitaihlgrance, Dec. 19, 1969,

Int. Cl. C07c 149/20 US. Cl. 260-481 R 1 Claim ABSTRACT OF THE DISCLOSURE The invention relates to a therapeutical composition useful in particular for the prevention and treatment of injuries due to ionizing radiations.

Said composition comprises, as active ingredient, a compound responding to either one of the following formulae: (A) (B) in which R is a straightor branched-chain alkyl group having 1-18 carbon atoms and R is hydrogen, COOH or COOR" in which R" is a straightor branched-chain alkyl group having 1-18 carbon atoms.

The invention relates also to a process for the preparation of the active ingredient of said composition and to the new compounds of Formula A or B.

This invention relates to a therapeutical composition useful in particular for the prevention and treatment of injuries due to ionizing radiations, to a process for the preparation of the active ingredient of this composition and to new condensation products of glyoxylic acid esters with aminothiols.

It is known that cysteamine is an excellent radio-protector; however, it has the drawback of a relatively high toxicity which detracts from its usefulness.

It is an object of this invention to provide a therapeutical composition useful for the prevention and treatment of injuries due to ionizing radiations, having an efliciency of the same order, if not higher, than that of cysteamine, while being substantially less toxic than the latter.

The composition according to the invention is characterized in that it comprises, as active ingredient, a compound responding to either one of the following formulae:

oif on-oooa or CH: \CHCOOR R' H-NH: )11 R- H n s s 0 orr-ooon or 015 \CHOOOR (lI-Ir-NHz H a'- 3 NE in which R is a straightor branched-chain alkyl group having 1-6 carbon atoms and R is hydrogen, COOH or COOR" in which R is a straightor branched-chain alkyl group having 1-6 carbon atoms.

3,790,623 Patented Feb. 5, 1974 ice It will be noted that compounds of Formula B are derived from compounds of Formula A by loss of one water molecule. The bases of Formula A are generally highly unstable and are converted to the corresponding bases of Formula B. In contrast, the addition salts (hydrochloride, oxalate, and the like) of bases of Formula A are stable and are preferred.

All the compounds forming the active ingredient of the composition according to the invention are new, except those in which, in Formula B, there are simultaneously R=CH or C H and R'=H, and, in this respect, they constitute one of the objects of the invention. The compounds already known were prepared by Z. N. Pazenko (Ukrain. Khim. Zhur., 1958, 24, 632) by methods entirely different from the process claimed here, and have never been de-- scribed as possessing any therapeutical activity.

The invention relates also to a process for the preparation of compounds of Formula A or B above, comprising reacting a compound of the formula:

With an alkyl glyoxylate of the formula OCHCOOR (D) or with a half-acetal-ester of the formula R OCHC O O R in the presence of an alcohol of the formula ROH, R and R having the above defined meanings, and collecting the resulting condensation product.

In the process according to the invention, the alkyl glyoxylate or half-acetal-ester is preferably used in a slight excess over the stoichiometric amount, for example in an amount of 1.1 mole per mole of compound of Formula C.

The compound of Formula C, which may be typically cysteamine, L-, D- or DL-cysteine, may be used as such or as the hydrochloride.

The reaction may be carried out in the cold or by boiling under refluxing conditions; it lasts generally several hours. The condensation product may be allowed tocrystallize on cooling, and then filtered and recrystallized from a suitable solvent. The solvent may also be removed by distillation in vacuo, and the residue taken up into a small amount of water and then made neutral with an alkali metal bicarbonate solution, after which the base may be extracted with ether; the residue from the evaporation of ether is then distilled in vacuo. The resulting base may be converted to the hydrochloride, using hydrochloric acid in methanol or ethanol solution.

The esters of thiazolidine-Z-carboxylic acid of Formula B in which R'=H may also be prepared by merely heating alkyl 2-(2-amino-ethylthio)glycolates of Formula A in an alcohol of formula ROH in which R is as previously defined.

The following non limiting examples are set forth to illustrate the process for the preparation of compounds of Formulae A and B. (Temperatures: C.)

Example 1.-Methyl 2-(Z-amino-ethylthio)-glycolate, hydrochloric acid salt (1) (Formula A; R-=CH,)

A solution of methyl glyoxylate (30 g.; 0.33 mole) in methanol (30 ml.) is added to cysteamine hydrochloride (34 g.; 0.03 mole) dissolved in methanol (150 ml.). After 3-5 hours of contact, the methanol is removed in vacuo, in a water bath at 40 C., by means of a rotary evaporator. The residue is recrystallized from ml. of methanol and is then dried in a desiccator, in vacuo, over phosphorus pentoxide. Slow M.P.=108-109 (dec.). Yield: 73%.

201.68: C, 29.77; H, 5.99; Cl, 17.58; N, 6.94; S, 15.89.

Found (percent): C, 29.50; H, 5.96; Cl, 17.61; N, 7.11; S, 15.74.

Example 2.Methyl 2-(Z-amino-ethylthio)glycolate, oxalic acid salt (II) Analysis.-Calculated (percent) for C H NO S) C H O C, 34.27; H, 5.75; N, 6.66. Found (percent): C, 34.06; H, 5.80; N, 7.24.

Example 3.Ethyl 2-(2-amino-ethylthio)glycoate, oxalic acid salt (III) ((Formula A; R=C H The compound is prepared in the same manner as Com pound II, substituting the methyl glyoxylate with ethyl glyoxylate. M.P'.=8l-82 (dec.).

Example 4.Propyl 2-(Z-amino-ethylthio)glycolate, oxalic acid salt (IV) (Formula A; R=C3H7) The compound is prepared in the same manner as oxalate (III), substituting the ethyl glyoxylate with propyl 2 propoxy glycolate (half acetal ester, Formula E; R=C3Hq). M.P.=98 (dec.).

Example 5.Butyl 2-(Z-amino-ethylthio)glycolate, oxalic acid salt (V) (Formula A; R=C H The compound is prepared in the same manner as Compound 1V, starting from the half-acetal-butyl ester (E, R:=C H M.P.: 102 (dec.). Yield: 61%.

Analysis.Calculated (percent) for (C H NO S) C H O C, 42.79; H, 7.19; N, 5.55. Found (percent): C, 43.19; H, 7.36; -N, 5.49.

Example 6.Pentyl 2-(2-amino-ethylthio)glycolate, oxalic acid salt (VI) (Formula A; R=C H The compound is prepared in the same manner as Compound IV, starting from the half-acetal-pentyl ester (E, R=C H M.P.=.9092 (dec.).

Example 7.Isopentyl 2-(2-amino-ethylthio)-glycolate,

axalic acid salt (VII) (Formula A;

The compound is prepared in the same manner as Compound IV, starting from the half-acetal-isopentyl ester (E, R=CH CH CH(CH M.P.=9798 (dec.).

Example 8.Decyl 2-(2-amino-ethylthio)glycolate, oxalic acid salt (VIII) (Formula A; R=C H The compound is prepared in the same manner as Compound IV, starting from the half-acetal-decyl ester (E, R=decy1). M.P.=99100 (dec.).

Example 9. Methyl Z-L-(Z-amino-Z-methoxycarbonylethylthio)glycolate, hydrochloric acid salt (IX) (Formula A; R=CH R'=COOCH A solution of methyl glyoxylate (5 g.; 0.055 mole) in methanol (5 ml.) is added to L-cysteine hydrochloride (7.9 g.; 0.05 mole) dissolved in methanol (25 ml.). After standing twenty-four hours, the methanol is removed in vacuo, in a water-bath at 30-35 C., by means of a rotary evaporator. The residual oil crystallizes rapidly. M.P.=170 (dec.).

Analysis.Calculated (percent) for 'C H ClNO' S: C,

32.37; H, 5.43; Cl, 13.64; N, 5.39; S, 12.34. Found (percent): C, 32.32; H, 5.09; Cl, 13.92; N, 5.55, S, 12.51.

4 g Example l0.--Methyl thiazolidine-Z-carboxylate (X) (Formula B; R=CH R'=I-I) Method A.Cysteamine hydrochloride (11.3 g.; 0.1 mole) or cysteamine base (7.7 g.; 0.1 mole) are dissolved in methanol (50 ml.) and methyl glyoxylate (9.7 g.; 0.11 mole) or methyl 2-methoxyglycolate (13.2 g.; 0.11 mole) are then added thereto. The reaction mixture is heated to boiling, under refluxing conditions, during four hours. The methanol is removed in vacuo, the material is then taken up into a small amount of Water, neutralized with an alkali metal bicarbonate solution, and rapidly extracted with ether. The ether solution is dried over anhydrous extracted with ether. The ether solution is driedover anhydrous sodium sulfate and the ether is distilled off. The base is distilled in vacuo, B.P. =l15. Yield: 60%.

The hydrochloride is prepared by dissolution of the base in hydrochloric acid (in ethanol solution) and precipitation with anhydrous ether. M.P.=156158 C.

Analysis.Calculated (percent) for C H ClNO S: C, 32.69; H, 5.48; CI, 19.30; N, 7.62; S, 17.46. Found (percent): C, 32.55; H, 5.57; Cl, 19.58; N, 7.73; S, 17.64.

Method B.Methyl 2-(Z-amino-ethylthio)glycolate, hydrochloric acid salt (I) (10.8 g.; 0.05 mole) is heated with methanol (30 ml.) during six hours, to boiling under refluxing conditions. After distillation of the solvent to dryness, the resulting methyl thiazolidin-2-carboxylate, hydrochloric acid salt, is recrystallized.

Example 11.Methyl L-4-carboxy-thiazolidine-Z-carboxylate (XI) (Formula B; R=CH R'=COOH) A solution of methyl glyoxylate (19.5 g.; 0.22 mole) in methanol (30 ml.) is added to a suspension of L- cysteine (24.2 g.; 0.2 mole) in boiling methanol (200 ml.) The mixture becomes homogeneous, after which the slightly pink clear solution crystallizes on cooling. 17 g. of substantially pure material are collected. M.P.=149-152 C. Yield: 45%. A further 10.7 g. of product, M.P. 143- 149, are obtained after concentration of the motherliquors. Total yield: 72%. The combined crystals are recrystallized from methanol. After desiccation in vacuo over phosphorus pentoxide at laboratory temperature, the pure methyl L-4-carboxy-thiazolidine-Z-carboxylate melts at 151-152 C.

Analysis.Calculated (percent) for C H NO S: C, 37.70; H, 4.75; N, 7.33; S, 16.74. Found (percent): C, 37.62; H, 5.21; N, 7.22; S, 16.88.

The following compounds may be prepared using the procedure described above for the preparation of methyl thiazolidine-Z-carboxylate (X).

Example 12.-Methy1 L-thiazolidine-2,4-dicarboxylate,

hydrochloric acid salt (XII) (Formula B;

R=CH R'=COOCH The condensation is effected from L-cysteine hydrochloride and methyl glyoxylate in methanol solution, by contact at room temperature, during several hours. The

reaction mixture is concentrated in vacuo, and anhydrous ether is added to precipitate the hydrochloride. Yield: 70%. On recrystallization from three volumes of methanol and after drying in vacuo over phosphorus pentoxide,.the compound melts at l82l83 C.

Analysis.-Calculated (percent) for C H ClNO S: C, 34.78; H, 5.00; Cl, 14.66; N, 5.79; S, 13.26. Found (percent): C, 34.63; H, 5.15; Cl, 14.62; N, 5.97; S, 13.25.

Example l3.-Ethyl thiazolidine-Z-carboxylate, and its hydrochloride (XIII) (Formula B; R=C I-I R"=H) The condensation is efiected from cysteamine or its hydrochloride and ethyl glyoxylate, heated under refluxing conditions, in ethanol.

Base: B.P. =128'-130 C. Hydrochloride: M.P. =97- Analysis-Calculated (percent) for C,H ',C1NO,S:- 'C; 36.45; H, 6.17; N, 7.08. Found (percent): C, 36.27; H, 6.07; N, 7.36.

Example 14.-Propyl thiazolidineicarboxylate, and its hydrochloride (XIV) (Formula B; R1=C H R'-==H) The condensation is effected from cysteamine or its hydrochloride and propyl glyoxylate, heated in'propanol.

Base: B.P. =145 C. Hydrochloride: M.P.=85-88 C.

Analysis.Calculated (percent) for CqH14CI-NO2S1 C, 39.71; H, 6.66; Cl, 16.75; N,"6';'62. Found (percent): C, 39.61; H, 6.71; C1,- 16.93;-N, 6.44.

Example 15,'Isopro'pyl thiazolidine 2 carboxylate (XV) (Formula B; R-=CH(CII3)2, R' =H) The condensation is efiected from cysteine (or its hydrochloride) "and isopropyl glyoxy'late in isopropanol.

6 Example 20.Isopentyl thiazolidine-Z-carboxylate (XX) (Formula B; R'I=H) B.P. =l48 C. Y Analysis.-Calculated (percent) for C9H17NOz C, 53.33;H, 8.43; N, 6.89; S,,15.77. Found (percent): C, 53.52; H, 8.59; N, 6.70; S, 15.40.

Example 2l.--Hexyl thiazolidine-Z-carboxylate (XXI) (Formula B; R=C H R =H) Analysis.--Calculated for C H NO S: C, 55.27; H, 8.81; N, 6.44; S, 14.75. Found (percent): C, 55.42; H, 9.10; N, 6.23; S, 14.28.

Example 22.-Decyl thiazolidine-Z-carboxylate, oxalic acid salt (XXII) (Formula B; R =C 2p R==H) M.P. =118-120 C. (dec.)

Analysis.Calculated (percent) for c ur Nos, C H O C, 52.87; H, 8.04; N, 3.85. Found (percent):

C, 52.74; H, 7.75; N, 3.82.

The results of toxicological, pharmacological and biological tests reported below show the activity of the compounds prepared by the process according'to the invention.

(1) Acute toxicity:

The approximate LD values in mice, on intravenous, intraperitoneal or peroral administration, are as follows:

Compounds I X XI XII XIII XIV XVI LDao g-l 575 000 270 320 150 I.p 1, 000 400 1, 000 200 000 000 600 Peros 1,000 1,000 1,000 1,000 1,000 1,000 1,000

B.P. =128130 C.

Analysis.-Calculated (percent) ,for C H NO S: C, 50.76, H, 7.99; N, 7.40; S, .16.95."'Found (percent): C, 50.76; H, 7.95; N, 7.24; S, 16.98. y

1 In the following examples the condensations are carried out as described for Compounds XIII-XVI, from cysteamine or its hydrochloride andalkyl glyoxylate' or the suitable half-acetal-ester, heated in the corresponding alcohol. I T

Example 17.Isobutyl thiazolidine-Z-ciarboxylate (XVII) (Formula B; R=CH CH(CH R'|=H) B.P. =l39 C. 1 s Analysis.Calculated (percent) for C H NO S: C,- 50.76; H,.7.99; N, 7.40;. S, 16.95.- Found (percent) C, -50.78;1H, 8.20; N, 7.33; S, 16.55.

' Exempt; 18. Sec butyl thiazolidine-2-carboxylate (XVIII) I Analysis-Calculated (percent) as C Hi NO szj C, 53.33;H, '8.42;"N,6.89-; S, 15.77. Found (percent): C, 53.30; H, 8.60; N, 6.80; S, 15.81.

(2) Radio-protective effects:

(A) In a first series of tests, experiments were carried out with each Compound I, X, XI and XII of the preceding examples, in the following manner:

(a) A first lot of six mice was subjected to X-ray irradiation sufficient so that all animals die within 30 days, 50% of the animals having died by the tenth day (850 roentgens, within about ten minutes). This first lot was administered intraperitoneally 10 ml./kg. of solvent for the compound. In all four cases, the sunvival time for. 50% of the animals (ST 50) was of ten days and no mouse survived by the 30th day.

(b) A second lot of five male mice was not subjected to irradiation but was administered intra-peritoneally the maximum dose tolerated (MDT) of the test compound, dissolved in 10 mL/kg. of solvent. 'In all four cases, the mice survived on the 30th day.

(c) A third lot of fourteen mice (seven male and seven female) was irradiated with the same X-ray dose as the first lot and was treated intra-peritoneally, in the same manner as the second lot, with the maximum dose tolerated (MDT) dissolved in 10 mL/kg. of solvent. The following data were recorded: total number of fatal issues, 50% survival time (ST 50), variation of this I 5 0% survival time with respect to that (ten days) of untreated irradiated mice, and number of mice surviving on the 30th day.

In the following table: Miglycol is a trade name for a mixture of C -C fatty acid triglycerides (Noble Hochst Chimie).

doses, respectively: 850r (=irradiation LD 50% )-1000r In another series of tests, compound (XVI) was ex perimented under analogous conditions (solvent=Miglycol). The results obtained with suchtests are sum; marized in following Table II.

TABLE no I Dose of Irradiation (XVI), rngJ Survival Average 1 kg., i.p. Volume rate on survival ST Dose Time, (10 min. prior injecte the 30th time 50 Number of animals per let (T) min. to irradiation (mL) 1,000 18 5 male 500 0. 25

a (B) The following supplementary tests were carried out mam v 2 1 with Compound I, the outstanding efliciency of which was 150% su-nv vnn TIME s'nso) tnxrnnssnn IN ins s s demonstrated above. 1 y Anfina1s having (1) 'LD in mice, by the intra-peritoneal route, was I Ref r e 7 received I determined in a more accurate manner'by the up and j t i down method (A. W. Kinbal, et al., Radiation Research, 20 30 1957, 7, 1) and was found to be from 850 to 965 mg./kg., 5

i.e., about 900 mg./ kg. Thus, the maximum dose tolerated, 5 used in the tests, was set at 450 mg./kg. 3

o n c n -6 (2) Investigation of the radio-protective ability p 5 Eight lots of ten mice each were administered intraperitoneally the maximum dose tolerated of compound, i'.e., 450 mg./kg., and were irradiated at the following (=irradiati0n LD 90%) 1l5Orl300r1450rl60 0r- 1750r and 1900r.

During the same time, reference animals were administered intra-peritoneally the same amount per kg. of weight, and were then irradiated.

The results obtained are set forth hereinafter, in Tables III and IV.

TABLE III SURVIVAL RATE AFTER 30 DAYS (SR 30) EXPRESSED As PERCENTAGE I 4 Animals having Reference received ,Itmay be .notedthatST SOis also" increased in the protected animals; f

3 Determination ofthe irradiationiD ili'srbt'eeti 7 animals.

(4) Calculation of the dose reducing factor.

The irradiation LD in unprotected animals being 857:, the following result is obtained:

To conclude, Compound I is ara dioj-protector at leastas' efficient as cysteamine, 'while being substantially' less toxic thanthe latter. Y

'I-hecomposition according to the invention maybe used for preventive tar-curativepurposesagainstthe dis: orders andinjuries caused by ionizing radiations, in par ticular during. radio-therapeutical treatment .of, tumors or leukemias, making it possible for the diseased system :tO. tolerate higher and, consequently, more efiicient doses ofradiatioiis;.'

The composition is administrableparenterally, rectally, orally or topically at dosages from mg"; to I g. of, active ingredient per day. 3 I Forparenteral, rectal or. oral administration, it isformulated typically in unit dosage fonrn iin'which the-active ingredientis combined with awelllmown vehicle or excipient suitable for the selected pharmaceutical form.

Included among the suitable vehicles or excipients, are, for example:

( 1) for tablets, coated tablets or .capsulesfiiactose, talc,

starch, magnesium stearate, with the conventional suitable additives;

(2) for injectable solutions:

; me s asa, a 1

(3) for suppositories; semi-synthetic glyceride s cocoa-butter, and'the like; H1 T (4) for ointments, creamy-lotions: petrolatum, lanolin,

etc.

TfIlliistrative of such imit dosage forms are tablets; coated tablets or fcapsules containing 100-500 ingl of active ing'redient, ampoules containing 1'5 mli'of injectable solution containing 10 wt. percent of active ingredient dis tilled jwater (ite 'f 100-1500 mg. per dose); and suppositories containing 100-500 mg. of activeii ingredient.

For topical administration, the composition may be formulated as ointments, creams or lotions containing,v 1-.5 percentfofiactive'ingredientitogetlier with a suitable excipient. "t' .3 A

sterile pyrogen-free water,

Having now described my invention, what I claim and References Cited dsire to Secure by Letters Patent is: March, Adv. Organic Chem, McGraw-Hill Inc. (1968),

1. A compound of the formula:

S Herman et al., C.A. 72, 67238p (1970). f, 5 Kameyama et al., C.A. 74, 124716R (1971).

HNH1 H LORRAINE A. WEINBERGER, Primary Examiner in which R is selected from the straightand branchedchain alkyl groups having 1-18 carbon atoms and R is TERAPANF? Asslstant Exammer selected from hydrogen, COOH and COOR" in 10 CL which R" is selected from the straightand branchedchain alkyl groups having 1-18 carbon atoms, and its 3 311 pharmaceutically acceptable acid addition salts. 

